小鼠肌肉卫星细胞的原代培养和鉴定

发布时间：2017-12-31 17:14

本文关键词：小鼠肌肉卫星细胞的原代培养和鉴定　出处：《中国人民解放军军医进修学院》2007年硕士论文　论文类型：学位论文

【摘要】： 目的建立可靠的小鼠肌肉卫星细胞原代培养方法及鉴定技术。方法取新生小鼠后肢肌获得细胞悬液，经Percoll分离，结合差速贴壁法提纯肌肉卫星细胞。倒置显微镜观察，，绘制细胞生长曲线，采用结蛋白、α-肌动蛋白免疫细胞化学方法鉴定肌肉卫星细胞。结果原代骨骼肌卫星细胞在开始分化前呈圆形，通过细胞分离、提纯后纯度为80％以上。在F10培养液内肌肉卫星细胞生长良好，1天后细胞逐渐贴壁生长，细胞较小，分布均匀，多呈圆形；3-6天增长旺盛；到7-10天可生长至单层成片约50％～70％；12-14天后肌肉卫星细胞增殖减缓。10-15天的卫星细胞结蛋白、肌动蛋白染色清晰。结论使用Percoll分离、差速贴壁法的培养可获得纯度较高的肌肉卫星细胞，而结蛋白、肌动蛋白免疫细胞化学染色是鉴定肌卫星细胞的有效方法。
[Abstract]:Objective to establish a reliable primary culture method and identification technique for mouse muscle satellite cells. Methods Cell suspensions were obtained from the hindlimb muscles of newborn mice. The muscle satellite cells were isolated by Percoll and purified by differential adhesion method. The cell growth curve was drawn by inverted microscope and desmin was used. Muscle satellite cells were identified by 伪-actin immunocytochemistry. Results the primary skeletal muscle satellite cells were round before differentiation. The purity of the satellite cells was more than 80% after isolation. The muscle satellite cells grew well in F10 medium. After one day, the cells gradually adhered to the wall, the cells were small, distributed evenly, and most of them were round. The growth was strong in 3-6 days. To 7-10 days can grow to monolayer, about 50%; After 12-14 days, the proliferation of muscle satellite cells slowed. 10-15 days after desmin, actin staining was clear. Conclusion High purity muscle satellite cells can be obtained by Percoll isolation and differential adherence method, and desmin, actin immunocytochemical staining is an effective method for the identification of muscle satellite cells.
【学位授予单位】：中国人民解放军军医进修学院
【学位级别】：硕士
【学位授予年份】：2007
【分类号】：R329.2

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